This invention relates to a method of transplanting hemopoietic stem cells, and more particularly to a novel method of transplanting hemopoietic stem cells by which donor cells may be well sustained without graft failure/rejection.
In the so-called modern bone marrow transplantation (BMT), an HLA-matched individual is recruited as the marrow donor and the recipient, i.e. patient, is placed on a full-fledged immunosuppressive regimen for preventing graft rejection in a pathogen-free environment until post-transplantation hemoimmunological recovery has taken place. In recent years, the source of pluripotent hemopoietic stem cells, the true subject of a BMT, has been expanding from the bone marrow of an HLA-matched individual or a partially HLA-mismatched individual, autologous bone marrow, autologous peripheral blood, allogeneic peripheral blood and cord blood. To keep abreast of this expansion, the transplantation of purified pluripotent hemopoietic stem cells available from such sources has become feasible and said BMT is now subsumed in the expanded concept of hemopoietic stem cell transplantation.
By now the bone marrow recipient has come to include patients with aplastic anemia, leukemia, certain hereditary diseases, and even solid tumors such as malignant lymphoma and carcinoma of the breast and, in fact, said hemopoietic stem cell transplantation is currently performed in a broad spectrum of diseases.
For detailed information on such hemopoietic stem cell transplantations, xe2x80x9cHemopoietic Stem Cell Transplantation, Its Foundation and Clinical Practicexe2x80x9d [Modern Medicine, Special Issue, 53, 2, 1998] can be consulted and the descriptions given there are incorporated in this specification by reference.
Furthermore, autoimmune diseases have come to be regarded as stem cell disorders in recent years [International Journal of Molecular Medicine, 1:5-16, 1998] and treatment of various autoimmune diseases by BMT, particularly allogeneic bone marrow transplantation (allo-BMT), is a focus of attention today.
Playing a central role in the recent diversification of BMT is allo-BMT. Particularly in keeping abreast of the expansion and improved coordination of bone marrow banks, the number of cases receiving unrelated donor-host BMT is on a steady increase and countermeasures to graft failure/rejection and graft-versus-host disease (GVHD) as well as the effect of transplantation are the current subjects of debate and study.
The object of this invention is to provide a novel method of transplanting hemopoietic stem cells which overcomes the long-standing problems associated with hemopoietic stem cell transplantation, particularly the problem of graft failure/rejection.
In accordance with this invention there is provided a method of transplanting hemopoietic stem cells which comprises subjecting a graft recipient to a radiation treatment using an effective exposure dose for hemopoietic stem cell transplantation in advance and then transplanting hemopoietic stem cells from a graft donor via the portal vein.
The present invention further provides said transplantation method further comprising administering hemopoietic stem cells intravenously following the portal venous administration of hemopoietic stem cells; said transplantation method wherein the radiation treatment is carried out by total body irradiation using two divided doses a day; and said transplantation method which is used for the treatment of autoimmune diseases.
The method of transplanting hemopoietic stem cells according to the present invention accomplishes the above-mentioned object. Particularly, by using the method of the present invention, the incidence of graft failure or rejection can be drastically reduced and the graft cells be well sustained, thus, allowing the transplant to express its intrinsic effect to achieve the s objectives: hemoimmunological recovery through normalization of hemopoiesis in the recipient and contribution to the treatment of chronic myelocytic leukemia (CML), acute myelocytic leukemia (AML), acute lymphocytic leukemia (ALL), malignant lymphoma, multiple myeloma, aplastic anemia gravis, myelodysplastic syndrome (MDS) and other hereditary diseases, treatment of autoimmune diseases, and gene therapy by the gene transfer technique.
The hemopoietic stem cells to be transplanted by the method of the present invention are not particularly restricted but may, for example, be those cells which are used in the conventional hemopoietic stem cell transplantation. More particularly, the cells may be bone marrow cells including hemopoietic stem cells, peripheral blood cells (particularly peripheral blood cells including hemopoietic stem cells as recruited by administration of a cytokine such as granulocyte colony-stimulating factor (G-CSF), cord blood cells, and mixtures of such cells. The typically preferred hemopoietic stem cells are bone marrow cells.
The donor of hemopoietic stem cells is not particularly restricted, either, but can be judiciously selected according to the criteria generally used when the transplantation of hemopoietic stem cells is considered appropriate to the recipient (patient).
The procedures for harvest and isolation of such hemopoietic stem cells are well known to those skilled in the art and not different from the procedures used in the conventional hemopoietic stem cell transplantation.
The hemopoietic stem cells which are particularly preferred for the purposes of the present invention may for example be bone marrow cells supplemented with about 1-2% of T cells from the standpoint of enhancing the graft rate without eliciting the onset of GVHD. More particularly, hemopoietic stem cells for transplantation are generally prepared by adding the usual anti-T cell antibody (e.g. a mixture of anti-CD3 antibody or anti-CD4 antibody with anti-CD8 antibody) to a cell population and then adding the complement to kill the cells (T cells) coupled to the anti-T cell antibody and thereby remove the T cells from the population or by adding anti-T cell antibody and removing the cells coupled to the anti-T cell antibody selectively by the magnetic bead method. The purification (isolation) of T cells can be made by removing erythrocytes from peripheral blood to provide mononuclear cells in the routine manner, adding said anti-T cell antibody to this cell population and selectively recovering the cells coupled to the anti-T cell antibody by the magnetic bead method or by adding the anti-T cell antibody conjugated with a fluorescent dye to the mononuclear cell population and recovering the T cells with an automatic fluorescent separation hardware. The above-mentioned preferred bone marrow cells for use in the present invention, that is to say bone marrow cells supplemented with a given concentration of T cells, can be prepared by mixing the T cells purified as above with the marrow cells depleted of T cells as above.
The most outstanding feature of the hemopoietic stem cell transplantation method of the present invention resides in two essential requirements, namely the radiation treatment using an effective dose for hemopoietic stem cell transplantation in advance and the subsequent portal venous administration of hemopoietic stem cells. As to the procedural details other than those two requirements, the conventional procedures for transplantation of hemopoietic stem cells apply.
For example, the hemopoietic stem cells to be administered into the portal vein can be prepared in a suitable dosage form (hereinafter referred to briefly as dosage form) in the same manner as the various pharmaceutical dosage forms containing cellular components of this type. Thus, except that they are administered into the portal vein, said hemopoietic stem cells can be used in the same manner as in the usual transplantation of hemopoietic stem cells. Where desired, said hemopoietic stem cells can be provided in the form of an injection.
In preparing the above dosage form, a variety of pharmaceutically acceptable carriers can be utilized. The carrier may be any of the carriers well known in the art. Furthermore, in preparing said dosage form, various infusions in common use today can also be employed. In addition, the dosage forms can be extemporaneously prepared from the donor in transplantation.
There is no particular limitation on the dose amount of said dosage form, either, but the dose amount conventionally used in the transplantation of hemopoietic stem cells can be employed. The preferred dosage may for example be about 3xc3x97108 marrow cells/kg or more.
It is important that the portal venous administration of said dosage form be carried out after the pre-transplantation irradiation.
The radiation treatment as the preparation for transplantation is carried out by irradiating the graft recipient with an effective radiation dose for hemopoietic stem cell transplantation. The xe2x80x9ceffective radiation dosexe2x80x9d is characterized as an exposure dose frustrating recovery of the recipient""s bone marrow cells (lethal dose) and is not particularly restricted as far as it is a medically acceptable dose which is generally used.
This radiation treatment can be carried out by total body irradiation (TBI) in the usual manner, preferably by the fractionated irradiation method using about 2 divided doses, for example a total radiation dose of about 10-12 Gy given in about 2 divided doses, particularly two fractional doses of 5-6 Gy each.
In one preferred embodiment of the present invention, hemopoietic stem cells are administered in a single dose from the portal vein, usually within about 24 hours of said irradiation. By this transplantation method, the graft failure or rejection of the donor-derived cells can be successfully avoided.
A still more preferred embodiment of the present invention comprises administering a supplemental dose of hemopoietic stem cells intravenously following the portal venous administration of hemopoietic stem cells. In accordance with this method, not only the graft failure or rejection of donor-derived cells can be more positively obviated but the radiation dose for use in the preparation for transplantation can be reduced.
Optionally, portal venous administration may be substituted for the above-mentioned intravenous administration for supplemental transplantation.
Inasmuch as it is performed after the initial portal venous administration of hemopoietic stem cells, the timing of this supplemental administration is not particularly restricted but this second administration is preferably carried out within about 5 days of the initial portal venous administration.
The procedure, as such, which can be used for said portal venous administration and for said intravenous administration of hemopoietic stem cells is well known to those skilled in the art. For example, the laparoscope-aided administration method which is per se known can be used for portal transplantation.
The above portal venous administration, preferably followed by said intravenous administration, after the radiation treatment according to the above-mentioned schedule results in a successful maintenance of the grafted hemopoietic stem cells in the recipient.
Unless the effect of the present invention is not compromised, the method of the present invention can be used in combination with the routine medical treatments given in the transplantation of hemopoietic stem cells as well as the chemotherapy with other drugs. The drugs which can be used in this concomitant chemotherapy include but are not limited to immunosuppressants such as cyclophosphamide (CY), cyclosporin A (CsA), methotrexate (MTX), and tacrolimus (FK506). Regarding the dosage and administration of such drugs, reference can be made to the corresponding information on the known (commercial) preparations.